During the final stages of differentiation in mammalian keratinizing epithelia, the nucleated cells of the stratum granulosum are transformed into a thin protective sheath, the stratum corneum. Impermeable, keratinized epithelia simultaneously protect against desiccation by retarding water loss and by preventing penetration of noxious environmental substances. However, dispite its general imperviousness, certain substances preferentially traverse the stratum corneum. We propose: (1) To identify those structural components of keratinizing epithelia which subserve the barrier function. Particular emphasis will be placed on membrane features revealed by freeze fracture, and quantitative changes in cell surface area and in ultracryotomy sections, respectively, since these methods circumvent fixation and embedding artefacts. (2) To clarify the pathways of percutaneous transport across normal and perturbed stratum corneum. Lipid-soluble substances with known high fluxes across that stratum corneum will be precipitated in transit, or will be visualized by soluble-tracer autoradiography of frozen-dried stratum corneum. (3) To identify the chemical substances responsible for barrier function in keratinizing epithelia. Hydrophobic intercellular lipids derived from extruded lamellar bodies are considered the principal water-tight seal of the stratum corneum. The composition of lipids isolated from the stratum corneum will be compared to those from fractions rich in lamellar bodies, thereby pinpointing those substances important for barrier function. To further verify their importance for the water barrier, we will measure evaporative water loss across monolayers composed of these lipids. Finally, (4) to study the molecular events which precipitate or accompany the sudden transformation of viable granular cells into nucleate cornified cells. The presence of active ion shifts will be sought utilizing cytochemical techniques for Na-K ATPase and measurement of transepithelial potential difference. Passive ion fluxes will be detected by cryoscopy, cytochemically (pyroantimonate precipitation) and by x-ray analysis.